Transcription is a process in which RNA is formed from the DNA. The RNA formed has a complementary sequence to the template DNA that directs its synthesis. Only adenine that incorporates the thymine in DNA replication, directs the incorporation of uracil in RNA formation.
RNA formation in transription:
Three kinds of RNA generate during transcription in prokaryotes:
- Transfer RNA (tRNA): it carries amino acids during protein synthesis
- Messenger RNA (mRNA): it has the message for protein synthesis
- Ribosomal RNA (rRNA): it is the component of the ribosome
- Several small nuclear RNAs: catalyzes the splicing of mRNA
The messenger RNA of bacteria is polycistronic which means it has transcribed coding information of adjacent genes.
Transcription is selective:
The entire DNA molecule is not expressed in transcription in prokaryotes. RNA is synthesized only for some selected regions of DNA. On certain other regions of DNA, there may not be any transcription at all. The product formed in transcription is known as the primary transcript. Primary RNA transcript undergoes certain alterations (splicing, terminal additions, base modification, etc.) commonly known as post transcriptional modifications, to make a functionally active RNA.
Enzymes involved in RNA synthesis:
Bacterial transcription in prokaryotes involves the DNA dependent RNA polymerase called RNA polymerase. Prokaryotes have a single RNA polymerase that translates all types of RNAs.
- RNA polymerase I: it synthesizes rRNA
- RNA polymerase II: it synthesizes mRNA
- RNA polymerase III: it synthesizes tRNA
RNA synthesis is performed by the enzyme RNA polymerase. The RNA complementary to the template DNA is formed by using nucleotides ATP, CTP, GTP, and UTP. These nucleotides have ribose than deoxyribose.
RNA synthesis also occurs in a 5’ to 3’ direction, just like DNA synthesis. The new nucleotides are added at the 3’ end of the growing chain and by the activity of RNA polymerase pyrophosphate is formed.
ATP, CTP, GTP, UTP → →→→→→→→→→→ RNA + Pyrophosphate (PPi)
The pyrophosphate is then hydrolyzed by pyrophosphatase into orthophosphate. This hydrolysis makes RNA formation irreversible, as if the pyrophosphate level becomes too high then RNA will be degraded by the reverse of the polymerase chain reaction.
Polypeptide chains in RNA polymerase:
There are five types of polypeptide chains in bacterial RNA polymerase: alpha α, beta β, beta prime β’, omega ꙍ and sigma σ. The core enzyme contains four polypeptide chains: 2 alpha, beta, beta prime, and omega.
Functions of polypeptide chains
The sigma factor does not have catalytic activity but it helps the core enzyme recognize the promoter. The binding of the sigma factor to the core enzyme forms a complex known as RNA polymerase holoenzyme which is a six-subunit complex. Transcription in prokaryotes starts with the holoenzyme only and synthesis of RNA is completed by the core enzyme after its initiation.
Alpha: it involves the assembly of the core enzymes, interacts with some regulatory factors, and helps in recognizing promotor
Beta: it binds to ribonucleotide substrates
Beta’: it has a binding site for DNA
Omega: it helps in stabilizing the beta’ subunit confirmation
Transcription process in prokaryotes:
Transcription in prokaryotes has three processes:
Initiation starts when the RNA polymerase holoenzyme binds to the promoter of the gene. On the promotor of bacteria, there are two characteristic sequences:
TTGACA: it is a six base sequence and is present about 35 base pairs before the starting point of transcription
TATAAT: it is about 10 base pairs upstream of the transcription starting site and is known as the Pribnow box.
These are known as the -10 and -35 site regions as these show their distance from the first transcribed nucleotide. RNA polymerase holoenzyme recognizes the specific sequences at the -10 and -35 sites of the promotor (all of the promotors) so the nucleotide sequence must be similar. These are called consensus sequences.
After binding to the promotor region, RNA polymerase unwinds the DNA without any help of helicase. Hydrogen bonds that keep DNA double stranded break easily as the -10 site is rich in adenine and thymine and this unwound DNA region is known as an open complex.
About 16 to 20 base pairs unwound region of DNA is known as the transcription bubble. During elongation, this bubble moves along the RNA polymerase as it transcribes the mRNA from the template DNA.
In this transcription bubble, an RNA-DNA hybrid is formed. While RNA polymerase moves from a 3’ to 5’ direction along with the DNA template, the sigma factor will be separated from core RNA polymerase and help another core enzyme to start transcription.
Core RNA polymerase form mRNA 5’ to 3’ direction, that is antiparallel or complementary to the template DNA. mRNA elongation continues and single stranded mRNA is released and behind the transcription bubble, two strands of DNA form their double helical structure.
Termination occurs when core RNA polymerase separated from template DNA. This happens in the terminator. There are nucleotides sequence before the prokaryotic terminator that form hydrogen bonds with single stranded RNA when transcribed into RNA. This pairing forms a hairpin loop structure and this structure stops RNA polymerase to transcribe DNA.
Terminators are of two types:
- Rho dependent
- Rho independent
Rho independent termination involves the six uridine residues which follow the mRNA hairpin. After the formation of a hairpin loop, the RNA polymerase stops, and the base pairs of A-U in uracil rich region of the terminator are not strong enough to hold the RNA-DNA duplex; then the RNA polymerase falls.
Rho dependent termination does not have the hairpin or a poly-uracil region. It has the rho factor that binds to mRNA and moves along it until it reaches RNA polymerase. Rho has helicase activity and causes RNA polymerase to separate from mRNA by unwinding the RNA-DNA complex.
Willey, J. M., Sandman, K. M., Wood, D. H., & Prescott, L. M. (2019). Prescott’s microbiology (11th ed.). McGraw Hill.